Optic-cup morphogenesis in vitro

One of the more landmark papers in Stem Cell Biology/Regenerative Medicine was published in Nature last week. A group of Japanese scientists managed to take Embryonic Stem cells (ES cells) in a three-dimensional culture medium and managed to produce an almost entire mouse retina (the optic cup). I’d like to provide a basic overview of the research and its potential implications.

The retina is the photosensitive tissue which lines the inner surface of your eye. When light falls on the retina, it initiates a cascade of reactions in the appropriate cells which result in nerve impulses that travel via the optic nerve to the brain and create the visual experiences that we have and mediate other light responses. The layers of the retina are shown in this diagram. Notice how the light enters from below and has to pass through a jungle of neurons before they actually reach the rods and the cones which are the actual photosensitive components of the retina. This sloppy design actually betrays the humble evolutionary origin of the structure where nature never had the chance to go back to the drawing board and redesign it altogether but had to build up on whatever inefficient infrastructure it had.


The complexity of the structure, as evident from the picture has always been of considerable interest to biologists. The phylogenetic roots of the structure has been a hot topic in creationist circles and it is probably due to the most popular quote-mine of all time in biology. Charles Darwin wrote in 1872,

To suppose that the eye, with all its inimitable contrivances for adjusting the focus to different distances, for admitting different amounts of light, and for the correction of spherical and chromatic aberration, could have been formed by natural selection, seems, I freely confess, absurd in the highest possible degree.

However, the part that follows soon is often left out!

Yet reason tells me, that if numerous gradations from a perfect and complex eye to one very imperfect and simple, each grade being useful to its possessor, can be shown to exist; if further, the eye does vary ever so slightly, and the variations be inherited, which is certainly the case; and if any variation or modification in the organ be ever useful to an animal under changing conditions of life, then the difficulty of believing that a perfect and complex eye could be formed by natural selection, though insuperable by our imagination, can hardly be considered real. How a nerve comes to be sensitive to light, hardly concerns us more than how life itself first originated; but I may remark that several facts make me suspect that any sensitive nerve may be rendered sensitive to light, and likewise to those coarser vibrations of the air which produce sound.

The phylogeny of the eye has now been researched in quite detail and no sane person now denies the claim that the eye evolved (a number of times) in different species across the animal kingdom. The ontogeny is still pretty much in the dark but in the aforementioned experiment, scientists have, for the first time, demonstrated that the organogenesis is more-or-less a self-organising process.

So how did they do it?

ES cells are pleuripotent cells meaning that they have the capacity to form almost any structure in the body. Under proper circumstances, these cells differentiate into progenitor cells appropriate for the development pathway involved. This is achieved by a complicated interplay of gene-expression and shaping of the epigenetic landscape. The process was previously thought irreversible but more recent experiments suggest that it can be reversed under certain experimental conditions. The gene-expression profile of the progenitor cells provide us with a tool to identify them. In this experiment, they tagged the genes which are specifically expressed in the retina with green fluorescent protein in order to track them.

So they took a culture medium (called the Serum free floating culture of the SFEB-q type) and cultured the ES cells in them. With the treatment of activin, retinal differentiation could be induced in this culture sans the epithelium. Further treatment with basement membrane matrix components led to the epithelial formation. A high percentage of the total cells also became positive for the retinal cell markers (as evident from their glowing due to the green-fluorescent protein tagging). A similar induction if retinal cells could also be achieved by treating the culture with a concoction of purified laminin and entactin.

What followed from there was more or less on autopilot mode with a few tweaking involved here and there. By the sixth day, the culture had separated into regions of retinal and other cell types. On day 7, the retinal cell aggregates showed formation of hemospherical epithelial vesicles evaginating from the main body with upto four vesicles per aggregrate. On days 8-10, the vesicles underwent a dynamic shape change and formed a two-walled cup-like structure. The distal portion of the epithelium progressively folded to give rise to something very similar to the optic cup in the embryo. It then exhibited interkinetic nuclear migration and subsequently generated stratified neural retinal tissue as seen in the live organism! The proximal portion differentiated into mechanically rigid pigment epithelium with a marker profile reminiscent of that of the retinal pigment epithelium progenitors.

The process is absolutely fantabulous to watch and you can do so by going to the Nature website and clicking on the “Movies” tab of the “Supplementary Information” page of the article. If not anything else, green fluorescent structures folding onto themselves is a pretty sight!

I have summarised the experiment but it hardly captures the awesomeness of the actual process. Interested readers are requested to read the original article for a more detailed account.

Now why is this experiment so important? Firstly, it greatly reduces the complexity of the organogenesis of a complicated structure like the eye. Secondly, it has a huge implication in the fields of regenerative medicine and tissue engineering where we might soon be able to grow human retinas in vitro for therapeutic and research purposes. It also demonstrates the power of self-directed processes in biology which often result in the emergence of unfathomable complexity and thus it instills the confidence in us to try and grow more complicated structures in the laboratory. Last but not the least, the process is damn interesting and if you don’t agree you f*** off!


– Debayan

(PS. This post was typed in haste and hence I have skipped over most of the important aspects of the research. If anyone has a query, drop by a comment and I’ll be more than happy to reply. )







4 responses to “Optic-cup morphogenesis in vitro

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